To mimic the action of imidaclothiz in environment more easily, pure strain were used to biotransform it. Hydroxyl imidaclothiz and olefin imidaclothiz are formed as the two main metabolites of imidaclothiz during its metabolism by Stenotrophomonas maltophilia. A new method has been developed for the simultaneous analysis of these metabolites by HPLC at 250 nm as a unimodal elution peak, even though wavelengths of 269, 250 and 314 nm are ideal for the individual analysis of imidaclothiz, hydroxyl imidaclothiz and olefin imidaclothiz, respectively. The solubilities of imidaclothiz, hydroxyl imidaclothiz and olefin imidaclothiz were determined to be 0.25, 0.9 and 1.15 g/l, respectively, by HPLC using a standard curve equation. Although hydroxyl imidaclothiz could be converted to olefin imidaclothiz under acidic conditions (e.g., HCl or H₂SO₄) it could not be transformed to olefin imidaclothiz by S. maltophilia. Consequently, we have proposed a partial metabolic pathway for imidaclothiz, which involves the simultaneous formation of olefin imidaclothiz and hydroxyl imidaclothiz. These results of pure biotransformation made it easier to understand the dynamic activities of insecticides in environment and will hopefully be referenced and support furthering studies on the metabolism of imidaclothiz in soil in future following application.